Identification and characterization of a target antigen of a monoclonal antibody directed against Eimeria tenella merozoites

Abstract

Monoclonal antibodies (Mab) were produced against Eimeria tenella merozoites. A single Mab, LPMC-61, was selected because of its ability to bind to merozoites by indirect immunofluorescence assay (IFA) and to inhibit in vitro sporozoite development. Mab LPMC-61 reacts with an approx. 10-12-kDa merozoite polypeptide in reduced SDS-PAGE, but with an approx. 80-kDa protein in non-reduced SDS-PAGE. The monoclonal recognizes similarly sized polypeptides in E. tenella sporozoites, oocysts and schizonts. A partial cDNA (LPMC-61f) encoding the LPMC-61 antigen was identified from an E. tenella sporozoite cDNA library in bacteriophage lambda gt11. In addition to Mab LPMC-61, the recombinant beta-galactosidase/LPMC-61f fusion protein is recognized by hyperimmune rabbit anti-E. tenella sporozoite serum, rabbit anti-E. tenella merozoite serum, and E. tenella-infected and immune chicken sera. DNA sequencing of LPMC-61f cDNA showed that the putative protein has an unusual tandem, non-perfect repeated sequence, with glutamine comprising about 48% of the predicted amino acids. A hydropathicity plot of the predicted amino acid sequence shows a central hydrophilic region, consisting of the repeated sequences, surrounded by hydrophobic regions on both sides. Since the merozoite stage of avian Eimeria has been implicated in the induction of a protective immune response in chickens, LPMC-61 may be an important immunogen for use as a vaccine against E. tenella.